Date Published: March 19, 2012
Publisher: Public Library of Science
Author(s): Pedro Eitz Ferrer, Paul Frederick, Jacqueline M. Gulbis, Grant Dewson, Ruth M. Kluck, Andrei L. Gartel. http://doi.org/10.1371/journal.pone.0031510
One of two proapoptotic Bcl-2 proteins, Bak or Bax, is required to permeabilize the mitochondrial outer membrane during apoptosis. While Bax is mostly cytosolic and translocates to mitochondria following an apoptotic stimulus, Bak is constitutively integrated within the outer membrane. Membrane anchorage occurs via a C-terminal transmembrane domain that has been studied in Bax but not in Bak, therefore what governs their distinct subcellular distribution is uncertain. In addition, whether the distinct subcellular distributions of Bak and Bax contributes to their differential regulation during apoptosis remains unclear.
To gain insight into Bak and Bax targeting to mitochondria, elements of the Bak C-terminus were mutated, or swapped with those of Bax. Truncation of the C-terminal six residues (C-segment) or substitution of three basic residues within the C-segment destabilized Bak. Replacing the Bak C-segment with that from Bax rescued stability and function, but unexpectedly resulted in a semi-cytosolic protein, termed Bak/BaxCS. When in the cytosol, both Bax and Bak/BaxCS sequestered their hydrophobic transmembrane domains in their hydrophobic surface groove. Upon apoptotic signalling, Bak/BaxCS translocated to the mitochondrial outer membrane, inserted its transmembrane domain, oligomerized, and released cytochrome c. Despite this Bax-like subcellular distribution, Bak/BaxCS retained Bak-like regulation following targeting of Mcl-1.
Residues in the C-segment of Bak and of Bax contribute to their distinct subcellular localizations. That a semi-cytosolic form of Bak, Bak/BaxCS, could translocate to mitochondria and release cytochrome c indicates that Bak and Bax share a conserved mode of activation. In addition, the differential regulation of Bak and Bax by Mcl-1 is predominantly independent of the initial subcellular localizations of Bak and Bax.
The intrinsic or mitochondrial pathway of apoptosis is regulated by the Bcl-2 protein family, with two members, Bak and Bax, required to permeabilize the mitochondrial outer membrane (OM) , . During OM permeabilization, Bak and Bax undergo significant conformation change involving exposure of N-terminal epitopes and homo-oligomerization , , , ,  to form an as yet undefined pore. Bak conformation change also involves transient exposure of the BH3 domain that then binds to the hydrophobic groove of another activated Bak molecule to form symmetric dimers , , , with the same process also evident for Bax , , , . Symmetric dimers of Bak and of Bax can then be linked by an α6∶α6 interface into higher order oligomers that likely constitute the apoptotic pore , .